The metabolism of isoleucine (Ile) is known to involve transamination to S(+)-Alpha-keto-Beta-methylvalerate (S-KMVA) followed by oxidative decarboxylation to S-Alpha-methylbutyryl CoA and eventually to propionyl CoA. However, S-KMVA is also converted to R(-) Alpha-keto-Beta-methylvalerate (R-KMVA) at an unknown site, giving rise by transamination to alloisoleucine (Ale). The rate of this conversion and the rate at which R-KMVA is decarboxylated (to R-Alpha-methylbutyryl CoA and eventually to butyryl CoA) are unknown, as are several other aspects of Ile metabolism, especially in patients receiving KMVA therapeutically. We propose to pursue this problem by primed continuous infusions of [1-13C] isoleucine, plus two forms of labelled alloisoleucine. Following preliminary studies of whole body Ile flux and oxidation rates in normal man, infusions of [1-13C] isoleucine, [1-14C] alloisoleucine, and [methyl-2H3] alloisoleucine will be conducted first in dogs and then in man. By measuring isotopic enrichment of Ile, Ale, S-KMVA, R-KMVA, and expired CO2, the 4 fluxes (S greater than CO2, S greater than R, R greater than S, and R greater than CO2) can be estimated. I.v. and nasogastric infusions will be compared to assess the role of the splanchnic bed in these reactions. The effect of dietary isoleucine restriction on these fluxes will be studies. Organ homogenates from dogs will be examined for their capacity to catalyze these reactions. Patients with high plasma levels of Ale and KMVA, owing to chronic R,S-KMVA administration, will be infused i.v. with these same isotopes. The time course of circulating levels of Ile, Ale, S-KMVA, and R-KMVA following single loads of Ile, Ale, S-KMVA, or R-KMVA will be measured for two hours in normal subjects. Plasma of children with branched-chain ketoaciduria, treated and untreated, will be analyzed for Ile, Ale, S-KMVA, and R-KMVA to determine the steady-state relationships between these compounds in the presence of BCKA dehydrogenase deficiency. The results should clarify several aspects of Ile metabolism.